Abeles, in a Special Report published in Chemical and Engineering News 61, 48 (1983) discussed generally a class of enzyme inactivators referred to as "suicide enzyme inactivators" or "mechanism-based inactivators". These inactivators, whether natural or man-made products, have a configuration that resembles the natural substrate of a natural enzyme, and therefore, are recognized by the active site of the enzyme. When the enzyme reacts with this recognizable configuration, the inactivator is modified chemically and thus converted into a compound that reacts with the enzyme leading to consequent enzyme inactivation, usually irreversible in nature. By acting on the suicide substrate (inactivator), the enzyme brings about its own destruction. The author postulates that development of inhibitors or inactivators of specific enzymes may be useful by playing an important role in the development of drugs that can interfere with normal enzyme activity. Where such enzymes have been identified as being implicated in a disease state, the use of such inhibitors or inactivators would be useful to modulate enzyme activity in the treatment or prevention of the given disease state in which the enzyme is implicated.
Research efforts have focused on the action or the mechanism of phospholipases, such as phospholipase A.sub.2, as being involved in the regulation of many important cellular functions. See Lapetina, Chapter 21 "Phospholipase", Annual Reports in Medicinal Chemistry 19, 213, Academic Press Inc. (1984).
Attendant goals of research into inactivation or inhibition of phospholipase enzyme activity can be taken inter alia, from Chang et al., Biochemical Pharmacology 36, 2429 (1987). See also Dennis Biotechnology 5, 1294 (1987). These commentaries provide a review of the structure and biochemistry of certain phospholipases and direct the reader to other articles where more exhaustive treatment on this class of hydrolytic enzymes may be found. The authors express the suggestion that phospholipase activation may represent a rate-limiting step in the whole process of lipid mediator synthesis which has been found to be implicated in the existence of several disease states or conditions. Specifically, phospholipase A.sub.2 (PLA.sub.2) is thought to be the enzyme responsible for release of free arachidonic acid from membrane phospholipids that is believed implicated in the control of prostaglandin, leukotriene and related eicosanoid biosynthesis in inflammatory, etc. states. They add that inhibition of PLA.sub.2 activity may offer an attractive therapeutic approach to the design of novel drugs for the treatment of such inflammation and other tissue injuries. The authors provide source and structure references for a large number of phospholipase A.sub.2 enzymes and discuss the mechanism of the activation of these enzymes. Finally, they provide an analysis of the consequence of such enzyme activities, and therefore, provide a basis for a goal of drug design researchers to produce new classes of drug entities that may be useful in the treatment or control of various disease states or conditions by mediation at the level of phospholipase or other hydrolytic enzyme activity.
Thus, after PLA.sub.2 becomes activated, it apparently mediates a variety of pathophysiological reactions probably via its products, lysophospholipids and arachidonic acid, and several potent biologically active substances such as prostaglandins, leukotrienes, etc. Also, when the phospholipid substrate contains an alkyl ether or plasmalogen function, the products of PLA.sub.2 action include lysoPAF (PAF=platelet activating factor) or analogues which when acetylated produce PAF and analogues which are also potent biologically active substances. These products or co-products of PLA.sub.2 activation are believed to be cytotoxic substances and have been implicated in several human inflammatory and allergic conditions. That and other data suggests that PLA.sub.2 is most likely involved in inflammation and tissue injury associated with various diseases.
Increased levels of PLA.sub.2 have also been found in rheumatoid arthritic conditions, psoriasis, pancreatitis and septic shock. Myocardial ischemia is also believed implicated by PLA.sub.2 activation. PLA.sub.2 may also be critically involved in lung pathophysiology, especially asthma.
All of this data suggests that an inhibition or inactivation of PLA.sub.2 may be a promising one-step approach to interfere with the production of the products or co-products of such activation that are implicated in various disease states, thus leading to the potential alleviation of the disease process.
A rather detailed characterization of phospholipase enzymes is also supplied by Dennis, Enzymes XVI, 307, Academic Press Inc. (1983). This article provides characterization data for and purification of several phospholipases, the kinetics and reactions of which these phospholipases are believed responsible, biological effects and identification of sequences and conformational structure suggesting active site(s).
Phospholipase A.sub.2 inhibition and modification by manoalide was the subject of U.S. Pat. Nos. 4,447,445 and 4,616,089. Manoalide is a marine natural product isolated from sponge and has been demonstrated to have anti-inflammatory activity in vivo. Research on this natural product implicated it to be acting on the level of phospholipase A.sub.2, and it has been shown to be an inhibitor of both cobra and bee venom phospholipase A.sub.2 (PLA.sub.2). See Lombardo et al., J. Biol. Chem. 260, 7234 (1985) and Deems et al., Biochem. Biophys. Acta. 917, 258 (1987). Research on synthetic analogues extending from that is described by Reynolds et al., J. Am. Chem. Soc. 110, 5172 (1988). These studies show that manoalide causes a time-dependent irreversible inactivation of PLA.sub.2 indicated by the modification of lysine residues. A disadvantage attending the use of manoalide is its nonspecific reactivity with a variety of proteins.
Encouraged by this goal to produce endogenous substances that could modulate hydrolytic enzyme activity, such as phospholipase, it is an object of the present invention to produce substances that can interfere with disease states or conditions via molecular interaction of specific hydrolytic enzyme activity on a suicidal inactivation or inhibitor mechanistic level. Based upon such research and study, using phospholipase A.sub.2 as a model, the present research focused on the design of novel hydrolytic enzyme inhibitors (inactivators) that function via recognition by the active site of such enzymes resulting in inhibition of enzyme functionality. Thus, the inhibitors hereof invite functional suicide of the enzymatic activity.